Epidemic giardiasis caused by a contaminated public water supply.

In the period November 1, 1985 to January 31, 1986, 703 cases of giardiasis were reported in Pittsfield, Massachusetts (population 50,265). The community obtained its water from two main reservoirs (A and B) and an auxiliary reservoir (C). Potable water was chlorinated but not filtered. The incidence of illness peaked approximately two weeks after the city began obtaining a major portion of its water from reservoir C, which had not been used for three years. The attack rate of giardiasis for residents of areas supplied by reservoir C was 14.3/1000, compared with 7.0/1000 in areas that received no water from reservoir C. A case-control study showed that persons with giardiasis were more likely to be older and to have drunk more municipal water than household controls. A community telephone survey indicated that over 3,800 people could have had diarrhea that might have been caused by Giardia, and 95 per cent of households were either using alternate sources of drinking water or boiling municipal water. Environmental studies identified Giardia cysts in the water of reservoir C. Cysts were also detected in the two other reservoirs supplying the city, but at lower concentrations. This investigation highlights the risk of giardiasis associated with unfiltered surface water systems.     

105Changes in Nitric Oxide Levels After Deep Dermal injury in the Female,Red Duroc Pig (FRDP)

Introduction : Hypertrophic scar is a devastating sequel to burns and other tangential skin injuries. It follows deep dermal injuries and does not occur after superficial injuries. Nitric oxide (NO) plays many important roles in wound healing from inflammation to scar remodeling. Studies have shown that expression of nitric oxide synthase and nitric oxide production are decreased in human hypertrophic scar. However little is known about NO involvement in the early stages of hypertrophic scarring, because of the lack of an animal model. It was recently reported that the female red Duroc pig (FRDP) makes thick scar, which is similar to human hypertrophic scar. We hypothesized that NO production in wounds on the female, red Duroc pig is similar to that of human hypertrophic scar and that NO involvement in deep wounds is different from that in superficial wounds. Methods : Superficial (0.015” to 0.030”) and deep (0.045” to 0.060”) wounds were created on the backs of four FRDPs. Biopsies were collected at weeks 1.5, 4, 8 and 21 post wounding including samples of uninjured skin. Nitric oxide levels were measured with the Griess reaction assay and normalized with tissue protein level. Results : Superficial wounds healed with an invisible scar whereas the deep wounds healed with scar resembling mild hypertrophic scar. The thickness of the scars from the deep wounds was significantly greater than uninjured skin and healed superficial wounds (p < 0.01). NO levels were increased at 1.5 weeks in deep wounds compared to superficial wounds and uninjured skin (p < 0.05). At 8 weeks, NO levels in deep wounds had returned to the level of uninjured tissue and superficial wounds. By 21 weeks, NO levels had decreased significantly when compared to superficial wounds (p < 0.01). There were no differences in NO levels between uninjured skin and superficial wounds at any time point (p > 0.05). Conclusions : NO production is similar in late, deep wounds on the female, red Duroc pig to that reported in the literature for human hypertrophic scar further validating this animal model. NO production is quite different after deep wounds as compared to superficial wounds in the FRDP. Early elevation in nitric oxide production might account for excessive inflammation in deep wounds that become thick scars in the FRDP. Nitric oxide regulators and effects at early stages of scar formation should be elucidated further and the FRDP appears to be a useful model.     

Enhanced immunogenicity of pneumococcal surface adhesin A by genetic fusion to cytokines and evaluation of protective immunity in mice

Immunization of mice with pneumococcal surface adhesin A (PsaA) emulsified in complete Freund's adjuvant (CFA) provides protection against systemic infection with Streptococcus pneumoniae. Because the use of CFA is not acceptable in humans, we sought to develop alternative means of enhancing the immunogenicity of protein antigens of potential use in pneumococcal vaccines. We designed a series of genetic constructs in which coding sequences for PsaA were linked to sequences encoding either murine interleukin-2 (mIL-2), mIL-4, or two copies of an immunostimulatory nonapeptide derived from mIL-1beta. The PsaA-cytokine constructs were cloned and expressed in Escherichia coli. Mice immunized twice with PsaA-IL-2, or PsaA-IL-4 responded with PsaA-specific antibody production comparable in magnitude to that of mice primed with PsaA in CFA and boosted with PsaA in incomplete Freund's adjuvant (PsaA-Adj). Antibodies elicited by PsaA-Adj were predominantly of the immunoglobulin G1 (IgG1) subclass, while PsaA-IL-2 and PsaA-IL-4 elicited substantial amounts of IgG2a in addition to IgG1. Mice immunized with PsaA-Adj or PsaA-IL-4 were partially protected against intraperitoneal challenge with virulent S. pneumoniae (30% overall survival beyond 15 days postchallenge). Mice immunized with PsaA and no adjuvant or PsaA-IL-2 exhibited 0 or 5% survival rates, respectively, following challenge. In contrast, mice immunized twice with capsular polysaccharide were 100% protected. The modest levels of protection seen in mice immunized with PsaA and its more immunogenic derivatives may be explained in part by the relative inaccessibility of antibody to PsaA on the surface of encapsulated S. pneumoniae.     

Diagnosis of Epstein-Barr virus infection in hairy leukoplakia by using nucleic acid hybridization and noninvasive techniques.

The presence of Epstein-Barr virus (EBV) DNA in the epithelial cells of oral hairy leukoplakia is the confirming criterion in the diagnosis of this lesion, which occurs mainly in persons infected by the human immunodeficiency virus. Because hairy leukoplakia often presages the development of the acquired immune deficiency syndrome, it is important that suspicious lesions be accurately diagnosed. Commonly, biopsy tissue is removed for detection of EBV DNA by in situ hybridization, but biopsy is contraindicated in some patients. This study evaluated filter and cytospin in situ hybridization, two noninvasive techniques that examine epithelial cells swabbed from the surfaces of the lesions, for their sensitivity in detecting EBV DNA. As compared with tissue in situ hybridization, the filter and cytospin techniques had sensitivities of 100 and 92%, respectively. We conclude that these two noninvasive techniques can provide the clinician with an accurate alternative to biopsy whenever this human immunodeficiency virus-associated lesion is suspected.     

Induction of HLA-DR expression on thyroid follicular cells by cytomegalovirus infection in vitro. Evidence for a dual mechanism of induction.

Cytomegalovirus (CMV) infection of primary cultures established from human thyroid nodular and normal (paranodular) tissues resulted in induction of human leukocyte antigen (HLA) DR expression on thyroid follicular cells (TFC), as detected by cell-surface immunofluorescence staining with monoclonal antibodies (MAb). Two distinct modalities of induction were observed. The first type occurred in cultures of normal tissue obtained from CMV-seropositive but not seronegative donors, was detected on 30% to 50% of the TFCs, even though the vast majority of these cells failed to show any morphologic or antigenic evidence of individual CMV infection, and was associated with production of gamma-interferon (gamma-IFN) in vitro. The induced molecules displayed the characteristic DR polypeptide profile on immunoprecipitation and electrophoretic analysis. These results demonstrate that CMV infection of normal thyroid cultures may induce DR expression on TFCs in the absence of pre-existing lymphoid infiltrates and suggest that the induction is the result of an in vitro response to CMV by previously sensitized immunocompetent cells present in these primary cultures. Such a response, associated with the release of gamma-IFN, would induce DR expression on neighboring uninfected cells. The second mode of induction occurred in all CMV-infected cultures, regardless of their tissue origin (nodular or normal) or the serologic status of the donors. Up to 50% of infected TFCs at a late stage of infection, having fully developed CMV antigen-positive intranuclear inclusions, also displayed the cell-surface DR-related determinant recognized by one of the four anti-DR MAbs used. This induction was restricted to TFCs, while CMV-infected fibroblastoid cells present in the monolayers were invariably negative. Induction by CMV of major histocompatibility class II antigens on human epithelial cells may have significant implications in the development of normal immune responses against local viral infection, the enhancement of alloimmune rejection of grafted organs, and the generation of organ-specific autoimmune responses.     

An analytical model for the dissolution of different particle size samples of Bioglass in TRIS-buffered solution

We analyzed the early stages of reactivity of three different particle size samples of Bioglass 45S5 and a bulk sample in TRIS-buffered solution at pH 8. Ion release, measured with ion-coupled plasma emission spectroscopy, and pH variations are reported. It was demonstrated that differences in the initial surface area influence the increase in pH, the rate of elemental release, and the rate of calcium phosphate reprecipitation. In particular, a thicker Ca/P layer was obtained on larger particles. The equilibrium value of Si in solution was independent of sample form and amount of sample dissolved, and was always close to the value observed when bulk silica is dissolved at pH 8. An analytical model is proposed for cation release, based on a two-step mechanism. It was found that the early stage of dissolution was nearly diffusion controlled for larger particles and bulk samples. The second stage was similar to a first-order homogeneous dissolution. The influence of sample surface area/solution volume ratio seemed to be more complex than that proposed in the early works presented in the literature. It is suggested that variation of surface area has a significant impact on the course of the dissolution.     

Transformation of rabbit arterial smooth muscle cells with Simian virus 40

Summary In vitro studies were carried out to induce viral transformation of vascular smooth muscle cells. Cultured rabbit arterial smooth muscle cells were infected with simian virus 40 (SV 40), and transformed cultures were produced that exhibit altered morphology, increased growth rate and plating efficiency, growth on semi-solid substrate, and chromosomal abnormalities. Nuclear SV 40 T-antigen was detected in all cells of these cultures. Muscle-specific actin was identified by a specific monoclonal antibody suggesting retention of smooth muscle cell characteristics by the transformed cells. Significant cytoplasmic lipid accumulation occurred in transformed cells incubated with -very low density lipoprotein, as revealed both by chemical analyses and Nile Red lipid staining of the cultures. The transformed smooth muscle cells grow permanently in cell culture. Our investigations show that arterial smooth muscle cells transformed with SV 40 virus exhibit altered phenotypic properties distinct from that of normal arterial smooth muscle cells.     

Chronic therapy with recombinant tumor necrosis factor-alpha in autoimmune NZB/NZW F1 mice

We studied the effects of recombinant murine tumor necrosis factor-alpha (TNF-alpha) on autoimmune disease in lupus-prone NZB/NZW F1 (B/W) mice. Treatment with TNF-alpha, begun after the onset of clinical disease, improved survival relative to control mice: at age 10 months, 92% of mice treated with TNF-alpha were alive compared with 42% of control mice (P less than 0.05). Administration of TNF-alpha delayed the progression of renal disease, but sustained therapy did not prevent the eventual development of severe nephritis. Despite the improvement in survival, treatment with TNF-alpha did not inhibit anti-dsDNA antibody production. However, it accelerated T lymphocytopenia and abolished natural killer cell activity. These observations suggest that TNF-alpha may retard murine lupus in B/W mice through effects on cellular rather than humoral mechanisms. Our findings also indicate that the beneficial effects of TNF-alpha cannot be sustained indefinitely by chronic therapy.     

Use of Nile red stain in the detection of cholesteryl ester accumulation in acid lipase-deficient fibroblasts

The fluorescent hydrophobic probe Nile red was used to distinguish between normal human fibroblasts and fibroblasts from individuals with a genetic deficiency in lysosomal acid lipase activity (Wolman's disease and cholesteryl ester storage disease). The fluorescence of Nile red-stained cultured mutant cells, indicative of neutral lipid accumulation, was intense when compared microscopically with normal fibroblasts. The cholesteryl ester accumulation in the acid lipase-deficient fibroblasts was demonstrated qualitatively and quantitatively when cellular lipid extracts were subjected to thin-layer chromatography, followed by Nile red plate treatment and fluorescence spectrometry scanning. These results demonstrate the utility of the Nile red stain to document cellular lipid overloading. The techniques are simple to perform and can effectively supplement the standard enzymatic analysis used in the diagnosis of acid lipase deficiency.